PROJECT SUMMARY Anterior cruciate ligament (ACL) rupture is a common injury on young active subjects that have long term debilitating consequence. Meta analysis studies have demonstrated that 50% of the subjects after ACL rupture generate posttraumatic osteoarthritis (PTOA) 10 to 20 years after injury regardless of their clinical treatment. Perpetuation of the inflammatory response to injury is hypothesized as an important factor in the process of joint degeneration, thus representing an attractive target for therapy. In general, the inflammation observed in PTOA is chronic, low-grade, mediated primarily by the innate immune system. Although several mechanisms have been identified that can activate the inflammatory response in vitro, there is little known on how the inflammatory response occurs in vivo, since we lack the technology to track inflammation in vivo. Current methods to assess joint inflammation in vivo are not tissue-specific like the analysis of inflammatory factors, or lack the biological specificity such as imaging. Molecular imaging represents a natural approach to address the pathogenesis of PTOA, since it can image inflammation processes at the molecular level with complete joint coverage. Our research aims to develop a new imaging technology to measure the low-grade inflammation in PTOA in vivo using mimetic peptides that specifically target the interaction of hyaluronic acid with the cell surface receptor CD44, which play important roles in inflammatory signaling. The proposed mimetic peptides have shown anti inflammatory effect in cancer and rheumatoid arthritis, but their value in OA has not been explored. The selected molecular targets are preserved cross-species, so our research has potential for clinical translation. To capture different pathways on the same subject we propose to use a multimodal imaging approach combining magnetic resonance imaging (MRI) and near infrared (NIR) imaging. We will conduct experiments to validate the contrast agents in vitro. We will assess in vivo the contrast agent pharmacokinetics at early and late stage of disease, and control-validate. Finally, we will test the contrast agents in a longitudinal study. The results of this proposal will provide a platform to elucidate the pathophysiology of PTOA, help identifying new therapeutic targets, and track therapy response.